Glucosensing capacity of rainbow trout telencephalon.

To assess the hypothesis of glucosensing systems present in fish telencephalon, we first demonstrated in rainbow trout, by in situ hybridisation, the presence of glucokinase (GK). Then, we assessed the response of glucosensing markers in rainbow trout telencephalon 6 hours after i.c.v. treatment with glucose or 2-deoxyglucose (inducing glucoprivation). We evaluated the response of parameters related to the mechanisms dependent on GK, liver X receptor (LXR), mitochondrial activity, sweet taste receptor and sodium-glucose linked transporter 1 (SGLT-1). We also assessed mRNA abundance of neuropeptides involved in the metabolic control of food intake (agouti-related protein, neuropeptide Y, pro-opiomelanocortin, and cocaine- and amphetamine-related transcript), as well as the abundance and phosphorylation status of proteins possibly involved in linking glucosensing with neuropeptide expression, such as protein kinase B (AkT), AMP-activated protein kinase (AMPK), mechanistic target of rapamycin and cAMP response element-binding protein (CREB). The responses obtained support the presence in the telencephalon of a glucosensing mechanism based on GK and maybe one based on LXR, although they do not support the presence of mechanisms dependent on mitochondrial activity and SGLT-1. The mechanism based on sweet taste receptor responded to glucose but in a converse way to that characterised previously in the hypothalamus. In general, systems responded only to glucose but not to glucoprivation. Neuropeptides did not respond to glucose or glucoprivation. By contrast, the presence of glucose activates Akt and inhibits AMPK, CREB and forkhead box01. This is the first study in any vertebrate species in which the response to glucose of putative glucosensing mechanisms is demonstrated in the telencephalon. Their role might relate to processes other than homeostatic control of food intake, such as the hedonic and reward system.

Organisation of the cerebellar nucleus of the dogfish, Scyliorhinus canicula L.: a light microscopic, immunocytochemical, and ultrastructural study.

Elasmobranchs possess a well-developed cerebellum with an associated cerebellar nucleus. To determine whether the organization of this nucleus is comparable with that of the deep cerebellar nuclei of mammals, we studied the dogfish cerebellar nucleus with light microscopic methods (Nissl stain, Golgi method, reduced silver stain, NADPH-diaphorase histochemistry and immunocytochemistry) and with electron microscopy. We found the dogfish cerebellar nucleus to consist of about 1,050 large neurons, the ratio of Purkinje cells to cerebellar nucleus neurons being about 17:1. Immunocytochemistry showed large glutamatergic neurons in the main portions of the nucleus and small glutamate- and/or alpha-aminobutyric acid (GABA)-immunoreactive cells in the subventricular region of the nucleus. Large glutamatergic neurons corresponded to bipolar or triangular cells revealed by Golgi methods. Application of horseradish peroxidase to the cerebellar cortex produced the labelling of beaded fibres of Purkinje cells in the cerebellar nucleus. Unlike in mammals, GABAergic innervation of the cerebellar nucleus was scare: Purkinje cell axon terminals in the cerebellar nucleus did not appear to be GABA-immunoreactive, most GABAergic fibres being found in the subventricular neuropile. Some fibres immunoreactive to serotonin and somatostatin were also observed in the subventricular neuropile of the cerebellar nucleus. Three neuron types were distinguished with electron microscopy (types A to C). Type A cells were abundant and smooth-surfaced, and appeared to correspond to Golgi-impregnated neurons and large glutamate-immunoreactive cells. Type B neurons were scarce and possessed dendrites covered by sessile or stalked spines. Type C neurons were small cells located mainly in the medialmost region of the nucleus and corresponded to subventricular glutamate- and GABA-immunoreactive cells. Six types of synaptic bouton were observed (types I to VI). The most abundant (type I boutons) made symmetrical contacts and appeared to correspond to Purkinje cell axons. Type I boutons were the only type observed on perikarya and initial axon segments of type A cells. Type IV and type V boutons made complex glomerular-like asymmetrical contacts with spines of type B cells. Type VI boutons appeared to correspond to peptidergic and/or monoaminergic axons. The functional significance of these results is discussed.

Chick/quail chimeras with partial cerebellar grafts: an analysis of the origin and migration of cerebellar cells.

Chick/quail chimeras with partial cerebellar grafts have been performed to obtain further information about the origin and migratory movements of cerebellar cortical neurons. The grafts were performed by exchanging between these two species a precise, small portion of the E2 cerebellar primordium, as defined in Martinez and Alvarado-Mallart (Eur. J. Neurosci. 1:549-560, 1989). All grafts were done unilaterally. The chimeric cerebella, fixed at various developmental stages, were analyzed in serial Feulgen-stained preparations to map the distribution of donor and host cells in the ependymal layer (considered to be reminiscent of the primary germinative neuroepithelium) and in the various cortical layers. In some of the oldest cases, we also used antiquail immunostaining to recognize quail cells. In the ependymal layer, it has been possible to conclude that each hemicerebellar primordium undergoes a morphogenetic rotation that changes its rostrocaudal axis to a rostromedio-caudolateral direction. However, important individual variations were observed among the chimeric embryos with respect to the ependymal area expected to be formed by donor cells. These variations cannot be explained solely on the basis of microsurgical procedure; however, they suggest the existence of important reciprocal interaction between host and grafted neuroepithelia. Therefore, it was not possible to draw a precise fate map of the E2 cerebellar primordium. Nevertheless, the dispersion of grafted cells in the cerebellar cortex, when compared to the real extent of the ependymal grafted area in each particular case, provided important data: (1) The external granular layer (EGL), the secondary germinative epithelium, seems not to originate exclusively from the "germinative trigone," as is usually considered the case. It emerges from a larger but restricted portion of the primary cerebellar matrix extending about the caudal fourth or third of the ventricular epithelium, as defined after its morphogenetic rotation. (2) The Purkinje cells (PCs) develop from all areas of the cerebellar epithelium. Although the distribution of donor PCs parallels the grafted ventricular layer mediolaterally, donor PCs extend more in the rostrocaudal dimension. The PC layer is formed mainly by donor cells in the lobules underlain by the grafted ependymal layer. However, donor PCs are also observed in cortical lobules surmounting the host ventricular layer. In these lobules, the donor PCs form clusters of various widths interrupting the host PCs. Reciprocally, clusters of host PCs are also found in the lobules formed mainly by donor PCs. The alternate small clusters of donor or host PCs are surrounded by Bergmann fibers of the other species' origin.(ABSTRACT TRUNCATED AT 400 WORDS)

GABAergic neuronal circuits in the cerebellum of the dogfish Scyliorhinus canicula (Elasmobranchs): an immunocytochemical study.

The presence of GABAergic cells and fibres in the corpus cerebelli and auricles of a dogfish, Scyliorhinus canicula, was studied with immunocytochemistry. In both locations, two types of GABA-immunoreactive (GABA-ir) neurons were observed: stellate cells in the molecular layer and Golgi cells in the granular layer. Stellate cell axons gave rise to numerous GABA-ir boutons distributed throughout the molecular layer and a smaller number of boutons which contacted Purkinje cell perikarya. No GABA-ir baskets around Purkinje cells were observed. Golgi cells of the granular layer gave rise to numerous GABA-ir boutons which were located around cerebellar glomeruli. Purkinje cell perikarya and their axon terminals in the cerebellar nucleus were not GABA-immunoreactive. These findings are discussed in terms of the phylogeny of cerebellar circuits.

A tract-tracing study of the central projections of the mesencephalic nucleus of the trigeminus in the guppy (Lebistes reticulatus, teleostei), with some observations on the descending trigeminal tract.

We studied the central projections of the mesencephalic nucleus of the trigeminal nerve (MesV) in the guppy (Lebistes reticulatus), after application of horseradish peroxidase or fluorescein dextran amine into the eye orbit. A small number (1 to 13) of large mesencephalic trigeminal neurons were solid labeled in the ipsilateral rostral mesencephalon. At the level of the trigeminal nerve entrance, the united process of each mesencephalic trigeminal cell bifurcates, giving rise to a peripheral branch that exits in the trigeminal nerve and a descending branch that runs caudally in a medial bundle separated from the descending trigeminal tract. This bundle passes close to the visceromotor nuclei of the medulla oblongata. Descending processes give rise to short collaterals to the descending nucleus of the trigeminus and the ventrolateral reticular area. Most MesV descending fibres terminate in this ventrolateral field at the transition of the medulla to the spinal cord, but one or two fibres could be followed to the C6 level, where they give rise to collaterals to the dorsal funicular nucleus. No collaterals directed to the trigeminal motor nucleus, the cerebellum, or the mesencephalic tegmentum were observed. These projections were also compared with those of the descending trigeminal tract.

An immunocytochemical and ultrastructural study of a specialized glial region of the medulla oblongata of the adult and juvenile grey mullet.

Electron microscopy together with glial fibrillary acidic protein (GFAP) and vimentin immunocytochemistry reveals the presence of a specialized glial region in the octavolateral area of the medulla oblongata of adult and juvenile grey mullets, Chelon labrosus. Glial cells, which can he characterized as ependymal and subependymal cells, originate in the walls of the lateral recess of the fourth ventricle. They contain numerous gliofilaments and are connected through frequent gap junctions. Electron microscopy also reveals the lack of nerve cell parikarya and processes in this region. Immunocytochemistry reveals that the glial cells are strongly GFAP-positive in both adults and juveniles, whereas vimentin is only detected in this region in juveniles. The meninges associated with this glial region contains connective fibres, formed of very thick collagen fibres, that arc metachromatic and PAS-positive under light microscopy. These findings strongly support a structural role for this medullary specialization. Differences between adults and juveniles in the distribution of GFAP- and vimentin-immunoreactive structures in other regions of the medulla oblongata are also reported.

GOLGI cells of the cerebellum of the dogfish, Scyliorhinus canicula (elasmobranchs): a GOLGI and ultrastructural study.

GOLGI cells of the dogfish cerebellum were studied using both GOLGI methods and electron microscopy. The arborization of their dendrites and axonal processes is limited to the granular layer, which in this species forms characteristic granular eminences. The dendrites are rather simple. However, the various axons of a GOLGI cell form a dense plexus of thin beaded processes whose complexity approaches that observed in the mammalian cerebellum. Electron microscopy shows that the GOLGI cells are medium-sized neurons whose processes contact cerebellar glomerulus of the granular layer. The GOLGI cell perikarya are also contacted by mossy fibres.

New structural aspects of the synaptic contacts on Purkinje cells in an elasmobranch cerebellum.

Nerve fibre contacts on Purkinje cell perikarya in the cerebellum of the small-spotted dogfish (Scyliorhinus canicula) were studied using the Cajal reduced silver technique, Golgi methods and electron microscopy. Silver staining revealed axons with thick swellings close to the base of Purkinje cells. Golgi methods demonstrated the presence of 'pincushions' of somatic spines on Purkinje cells. Electron microscopy revealed flattened fibres that formed extensive synaptic contacts with the Purkinje cell 'pincushions'. It is proposed, on the basis of the ultrastructural features, that these fibres are climbing fibres. Their possible significance in terms of the evolution of cerebellar circuitry is discussed.